PROJECT SUMMARY/ABSTRACT Chronic liver disease and liver cancer are significant sources of morbidity and mortality, accounting for an estimated 2 million deaths annually. Non-alcoholic fatty liver disease (NAFLD) is a pertinent public health threat because it can lead to impaired metabolic function, cirrhosis, and primary liver cancer, and it affects up to 25% of U.S. adults. Our laboratory utilizes chemical and genetic modulation of liver development and adult liver function in zebrafish to identify signaling pathways that regulate liver growth, homeostasis, and disease. We have found that Vitamin D Receptor (Vdr) activity during embryonic liver development is essential for normal liver growth, and that Vdr stimulation modulates transcription of lipid metabolic effectors during early liver formation. Impaired VDR function is associated with a NAFLD, liver fibrosis, cirrhosis, and liver cancer in humans and animal models, but the functions of VDR signaling in liver development and disease are unknown. Our objective is to characterize the cellular and molecular functions of Vdr activity in zebrafish liver development and to evaluate the functions of Vdr in adult liver homeostasis and metabolic stress. In Aim 1, we will utilize chemical and genetic modulation of Vdr activity to define the impact of Vdr signaling on liver progenitor expansion and liver outgrowth. We will investigate mutant lines to characterize the consequence of vdr deficiency and impaired vitamin D metabolism on liver growth. Based on preliminary RNA-seq data, we will investigate the functions of PI3K/mTOR signaling and other candidate signaling pathways in vitamin D induction of liver growth. These experiments will uncover the impacts of Vdr signaling during liver formation. In Aim 2, we will characterize the functions of Vdr activity during adult liver homeostasis using histological and transcriptomic analysis, with a focus on hepatic fat accumulation and lipid metabolism. In addition, we will determine the impact of hepatocyte-specific Vdr impairment in zebrafish subjected to a high-fat high- cholesterol diet challenge. Since Vdr impairment is associated with NAFLD, we anticipate that Vdr impairment will sensitize adult fish to NAFLD pathogenesis following a high-fat high-cholesterol diet challenge. This work will provide detailed mechanistic insight into the functions of VDR signaling in liver development and disease and will illuminate the potential of vitamin D/VDR modulation as a strategy to prevent and/or treat chronic liver disease.